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Histopathological Effect of (Peganum harmala) Alcoholic Extract of Seeds on Liver and Kidney in Mice

Manal H.AL-Jborrey*, Ayyad W. Al-Shahwany

College of science, University of Baghdad, Iraq

Abstract: Toxicity still a global problem for the environment, agriculture and ultimately human health. The purpose of the study was to examine toxicity effects of Peganum harmala herbal extract on liver and kidney of albino Swiss mice. The herbal alcoholic extract used in different doses (1000, 2000, and 3000 mg/kg b.w) for 2 weeks. The liver and kidneys dissected out and quickly fixed in 10% formal saline for histologic study. The result indicated that the treated sections of the liver showed congestion of central vein surround the hepatic lobules with sever infiltration of inflammatory cells. In addition, the Kidneys showed congestion of blood vessels with enlargement of glomerular tuft and mild-severe infiltrate of chronic inflammatory cells compared to the control. The findings indicated that the administration of Peganum harmala extract has some adverse effects on the liver and kidneys of adult mice. The study showed that alcoholic herbal extract exposure might cause harmful effects to no target organisms, including humans. Finally, further studies may be carried out to examine toxicity effects for other herbal extract to induce the safety dose dependent to histopathological changes in the liver and kidney tissues of exposed mice.

Keywords: Peganum harmala, alcoholic extract, toxicity plant, Histopathological change

Citation: Manal H.AL-Jborrey, Ayyad W. Al-Shahwany, Histopathological Effect of (Peganum harmala) Alcoholic Extract of Seeds on Liver and Kidney in Mice. Biomedicine and Biotechnology, 5 (1). 1-5. January 2017. doi:10.12691/bb-5-1-1

1. Introduction

The plants considered as the most important natural ingredients found in the living environment and because these plants are important in economic terms, but the medical significance now become much more than that. These herbs according to the active ingredients which became as treasure for all researchers and they are looking for discover the benefit of these herb. There are large number of studies in this area have been made clear many secrets of medicinal and therapeutic benefits of these herb which consider as cheap drug and useful to treat many symptoms and disorder via using single herb [1]. Peganum harmala herb is one of medicinal plants in herbal medicine. It has different named according to place grow in like "harmal" in Saudi Arabia, "African Rue" in African and Australian regions. Peganum harmala L. (Syrian rue) a wild herb growing belong to the family Zygophyllaceae. In Iraq, the herb grow in north of Iraq especially in shaqllawa [2]; In Egypt, it grows wildly in the coastal region from Sallum to Rafa. It is claim to be an important medicinal plant report to be widely used in treatment of several diseases and symptoms as antithelmintic and protozoacidal agent, asthma and malaria. The herb has pharmacological and biological effect according to Kurata and Liu et al. [3,4] that P. harmala activities such as antibacterial and antifungal [5]. P. harmala contain different chemical compounds including flavinoids, amino acids, polysaccharids, volatile compounds and several kinds of alkaloids compounds and the vital of effective ingredients "alkaloid" which has the majority of the therapeutic properties of the harmala herb [6].

The aim of this study is to detect the histopath changes exhibited by toxicity of P. harmala seeds extract on living organs, particularly the liver and kidney.

2. Materials and Methods

2.1. Extract Preparation

A quantity of 100g of P. harmala seed was bought from a local market, grinded and extracted with (500 ml) of methanol for 2 days by Soxhelt apparatus at 40-60°C. The extract was filtered, evaporated to dryness at 40°C and stored until use. A small amount of extract obtained which was 25 g from the original weight). This crude extract was used to prepare the required doses (1000, 2000 and 3000 mg / kg b.w) to discover the lethality of high dose [7].

2.2. Experimental Design

In these studies, Albino mice were weighting 25-40 g. The animals were divided into four groups designated as A, B, C and D. Each group consisted of five animals treated with alcoholic extract of P. harmala for two weeks orally as follows:

Group A: treated with distilled water (control).

Group B: treated with 1000 mg / kg b.w of alcoholic extract of P. harmala for 2 weeks orally by gastric lavage.

Group C: treated with 2000 mg/kg b.w of alcoholic extract of P. harmala for 2 weeks orally by gastric lavage.

Group D: treated with 3000 mg/kg b.w of alcoholic extract of P. harmala for 2 weeks orally by gastric lavage.

2.3. Histological Study

Organs(liver and kidney)which were fixed in formalin (10%) for a few days were processed for sectioning then embedded in paraffin blocks and sections about (5μ)thick by using rotary microtome. The sections stained with Heamatoxylin and Eosin using routine procedure. The slide examined microscopically for observing the pathological changes [8].

3. Result

3.1. Effect of the Alcoholic Extract of (P. harmala) Seeds on Mice Body Weight

The results in Table 1 show significant differences in body weights compared with control.The increase in body weight of was in control treatment, which traded with water only. However, the other treatments were significant decreased the body weight specially (3000 mg/kg b.w).

Table 1. Mean ± S.D body weight (mg) of control group compared to the treated groups (1000, 2000 and 3000mg/kg) of alcoholic extract of (Peganum harmala)

4. Histological Study

4.1. The Liver

The microscopically screening of the control group liver appearance the section reveal radial arrangement of hepatocyte, sinusoid originates at lobules margin toward the hepatic vein c.v. as in Figure 1.

Figure 1. Liver section of group A (control) with normal architecture (H&E stain X40)

The liver of animal group treated with 1000mg/kg b.w of Peganum harmala extract the section show infiltration of few inflammatory cells as in Figure 2.

Figure 2. Liver section of group B treated with 1000 mg/kg of P. harmala extract (H&E 40X)

The liver section of group treated with dose 2000 mg/kg of Peganum harmala alcoholic extract showed enlargement of hepatocyte and presence of mitotic figure, there are few number of inflammatory cells infiltrated and congestion of blood vessels as in Figure 3.

Figure 3. Liver section of group C treated with 2000 mg/kg of P. harmala extract showing mitotic figure presence (H&E 40X)

The liver section of group treated with 3000mg/kg b.w of Peganum harmala extract showed congestion of central blood vessels with infiltration of inflammatory cells in Figure 4.

Figure 4. Liver section for the group D treated With 3000 mg/kg of P. harmala Extract showing congestion (H &E 40X)

5. The kidney

In kidney section of control group showing proximal convoluted tubules with different size together with renal glomerular appear as rounded structure surrounded by narrow Bowmans space as in Figure 5.

Figure 5. Microscopic section of kidney of control group with typical parenchyma (H & E stain X 40)

Kidney section of group treated with 1000mg/kg of Peganum harmala alcoholic extract revealed congestion of glumerular tuft with infiltration of inflammatory cells as in Figure 6.

Figure 6. kidney section for the group B treated with 1000 mg/kg of P. harmala extract ( H&E 40X)

The kidney section of group treated with 2000mg/kg b.w of P. harmala extract show sever hemorrhage of glumeruli with few number of inflammatory cells infiltrated in Figure 7.

Figure 7. kidney section for the group C treated with 2000 mg/kg of P. harmala extract showing hemorrhage of glomerulur tuft (H&E 40X)

The kidney section of animal group treated with 3000 mg/kg b.w of P. harmala extract showed hemorrhage and enlargement of congested glumerular tuft and focal aggregation of inflammatory cells (lymphocyte &neutrophyle) as in Figure 8.

Figure 8. kidney section for the group D treated with 3000 mg/kg of Peganum harmala extract showing congestion of glomerulur tuft (H&E 40X)

6. Discussion

In recent years, the seeds of herb (P. harmala ) has gained greater importance in the East region of world because the plants have numerous relation with religious and intellectual beliefs as well as the herb has distinctive smell which help leisure of life. Harmala herbs like many medicinal herbs, which have therapeutic at one dose and toxic at another [9]. According to Harmala herb it differ from other herb which there are little studies concerning about the effect of the plant unaccompanied on liver and kidney. Sub-acute toxicity helps to determine the nature toxic of herbal extract through evaluated the changes in body weight and physiological change in vital of organs. It is important to conclude the food intake during the safety of herbal extract with medicinal purpose. Therefore, the fewer intakes are reflecting on the physiological status of the animal and considered as best possible response to the active compounds presence in herb [10]. However, the study gave tried to investigate the effect of low and high doses of (P. harmala ) on these vital organs. Grossly, the decreasing in body weight after treated with (1000 ,2000 and 3000) mg/kg, activity to reduce cholesterol, triglycerides and LDL cholesterol and the mechanism efficacy of diverse medicinal herbs to reduce serum cholesterol level. This result might be due to the presence of different levels of alkaloid harmine, harmaline, and harmol present in P. harmala [11]. These alkaloids reported to have hypoglycemic properties [12]. In addition, alkaloids presence may be leads to same hepatotoxicity in liver of animal [13]. In other way, the results mean herbal extract of P. harmala have harmful effects, especially in high doses. This indicate that, there were interruption in the metabolism of carbohydrate, protein and fat after a period of treated the animals with herbal extract [14]. The changed in body weight may be reflect to the toxicity or marker to adverse effect of toxic ingredient existence in herbal extract [15]. The pathological changes in real organ "liver and kidney" which may be indicated to the injury or damage caused by toxicant [16]. In real study, the histopathological changes in liver showed some negative effects appear through cellular hypertrophy, which attributed to liver injuries or due to accumulation of lipid droplets or glycogen in the cytoplasm of hepatocyte and increased cytoplasmic organelles [17]. It appears from the previous that the liver possesses no table mitotic potentialities, which can be stimulated under a wide variety of conditions. The change in real organ may combined with loss of defect parenchyma then following removal parts of the tissue leading to necrobiosis necessitating repair that mean the mechanism of the mitotic reaction may be occur in livers with or without loss of hepatic tissue [18]. According to these considerations, it inclined to assume that the mitotic reaction in the livers of rats treated with herbal extract of P. harmala was the consequence of primary changes in the organ. This would mean that increased cell multiplication accompanied with loss of parenchyma, but the changes does not affect morphologic arrangement and that probably because the mitotic reaction. This dependent upon the amount of hepatic tissue lost. The real changes, which affect functional part of real organ or tissue, can be associated with variations in animal weight that had been observed [19]. Mild inflammation in liver tissue can observe with infiltration of inflammatory cell around the central vein region and sinusoidal spaces that contributed to hepatic oxidative stress [20,21]. The presence of focal inflammation was the most frequently seen in major inflammatory lesion in rodent toxicity studies which considered as chronic form of inflammation the primarily cells. It seen that, lymphocytes in the majority point to active inflammation predominately existence. In other side, there was significant number of neutrophils. The Both cells chiefly companied with macrophages cells [22]. Moreover, congestion of central blood vessel leads to hypoxia and because of oxygen and nutrient deprivation hepatocyte degenerate or eventually may undergo necrosis [23]. The increasing severity of inflammation in the liver of treated mice may due to augment lipid peroxidation, and to create the free radicals effects [24]. Failure of body to excrete may be due to the toxic material in herbal extract which caused glumerular damage, and that pathological changes impaired renal function that interfere with excretory function caused with decrease in blood osmotic pressure and subsequent decrease in drainage of tissue fluid which reflected through congestion the blood vessels of affected tissue [25]. In other hand, exposure the lab animal "mice" to herbal extract with different doses obviously affected interstitial space of glomeruli and tubules which seen the Glomeruli tuft appeared swollen of Bowman's spaces [26]. That primarily due to congestion of glomerular capillaries and thickening of glomerular basement membranes with mild proliferation of the glomerular epithelial cells and thickening of the Bowman's capsule and periglomerular fibrosis [26]. The result were recognized reduction in the size of the glomerular capillary tufts, dilatation of Bowman’s capsule, degeneration of the proximal tubular epithelial cell lining with loose of the apical microvilli with accumulation of exudates within the dilated tubular Lumina of both convoluted tubules [27,28,29]. The result shows the extensive degeneration of the tubular epithelium and glomerular damage. The kidney of group of herbal extracts treated show damaged capillary loops with glomeruli and tubules haemorrhage and that considered the primary effect which can see mainly distended to dilatation of proximal and distal tubules in the cortex of glumeruli. While secondary effect associated with inflammatory processes [30,31,32].

7. Conclusion

Sub-acute treatments with high doses of P. harmala extracts cause reduction in body weight gain and damage liver & kidney for lab animal. Further studies are required to isolate and identify the active constituents of the other medicinal plants to clarify the mechanism of their toxic effect.

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